Exposure to substances in the environment and the diet that disrupt endocrine signaling has dramatically increased in recent decades. While the effects of estrogenic compounds on the function of the reproductive system has been well studied, much less is known about how other organs, particularly the hematopoietic system, are impacted. Definitive hematopoietic stem cells (HSCs) both self-renew and differentiate to produce each of the mature blood cell lineages for the entire lifespan of the individual. Defects in HSC formation or differentiation, as a result of inherited or acquired genetic aberrations, can lead to bone marrow failure and leukemia. Most genes involved in HSC formation are highly conserved across vertebrates, and continue to regulate HSC self-renewal and differentiation in the adult. The transcription factor RUNX1, a frequent target of chromosomal mutation in leukemia, is expressed in all sites of de novo hematopoiesis and is absolutely required for HSC induction in mammals; runx1 expression is conserved in zebra fish. Through a zebra fish chemical screen for modifiers of runx1 expression, the principal investigators identified compounds that modulate estrogen signaling as potential regulators of HSC formation. They hypothesize that environmental estrogens exert an effect on HSC development through their estrogenic function and that exposure to these compounds could have long-lasting negative effects. Their long-term goal is to understand the molecular and cellular mechanisms by which environmental estrogens affect hematopoiesis. The objective here is to characterize the functional implications and vulnerable periods of environmental estrogen exposure on HSC formation, proliferation and function during development and recovery after injury, in both zebra fish and mice. The central hypothesis is that estrogenic compounds impact HSC formation during multiple distinct periods during development, affecting the primitive wave of erythropoiesis, the formation of the vascular hematopoietic niche and HSC proliferation. This hypothesis has been derived from their screening results and subsequent evaluation of the different time periods contributing to HSC formation. The rationale for the work is that a detailed understanding of the impact of environmental estrogens on blood stem cell growth and proliferation will enable a deeper understanding of the consequences of exposure to environmental estrogenic compounds beyond their action on reproductive organs. In Specific Aim 1, they will investigate the effects of estrogen signaling on HSC specification and proliferation during hematopoietic development and recovery from injury in zebra fish. In particular, they will identify the cellular and molecular targets of estrogen action and the developmental stages during which the organism is particularly vulnerable to its effects. In Specific Aim 2, they will analyze the specific effects of xenoestrogens (bisphenol A), dietary phytoestrogens (genistein), and mycoestrogens (zearalenone) during HSC formation; they will demonstrate that these effects are mediated by specific activation of estrogen signaling. Using murine assays, they will examine how these compounds effect HSC function across vertebrate species. PUBLIC HEALTH RELEVANCE: Hematopoietic stem cells form the foundation of our blood and immune system; the formation and function of these cells are carefully controlled in the body. The proposed research will help to identify environmental factors that impact the birth and propagation of these stem cells. This work has great relevance for the identification of estrogenic compounds that lead to leukemia, and for recovery from bone marrow transplants.